IPCS INCHEM Home

    PROTEASE (Streptomyces fradiae)

    Explanation

         This enzyme preparation has not been previously reviewed by the
    Joint FAO/WHO Expert Committee on Food Additives.

         Each production batch is commenced with a selected colony of
    S. fradiae (strain No. 2019 in the mycological collection of the
    Museum National d'Histoire Naturelle, Laboratoire de Cryptogamie,
    Paris). The incubation medium in the industrial fermenter in the final
    stage (approximately 60 hours) of mycelium production is (wt/vol%):
    soya flour, 1; corn starch, 3; corn steep liquour, 0.5; K2HPO4, 0.1;
    Fe2(SO4)3, 0.0015; MgSO4, 0.005; CaCl2, 0.05. The medium is
    filtered, concentrated and protein is salted out. Crude protease is
    redissolved and processed with further salting out to yield "pure"
    (i.e., concentrated) protease, with an activity of about 12 000 Anson
    Units/mg. The diluted (industrial) enzyme is prepared by mixing with
    lactose to give a final activity of 1000 Anson Units/mg. The
    industrial product is free of amylase, invertase and lipase activity.
    A solution of 100 mg/ml of the industrial product had no antibiotic
    activity against a battery of test organisms.

    BIOLOGICAL DATA

    TOXICOLOGICAL STUDIES

    Short-term studies

    Rat

         Wistar rats distributed in 3 groups each containing 12 males and
    12 females were fed S. fradiae protease at levels of 0, 0.1 and 0.2%
    in the diet for 185 days. The treated diets contained 2 million and 4
    million Anson Units of enzyme activity, respectively, per kilogram of
    foodstuff. There were no significant differences between groups in
    growth rate or food intake. One male control rat died with a
    pancreatic tumour. One female rat in the 0.2% test group had a gastric
    lesion that was revealed in routine post mortem examination at the end
    of the trial period. No other lesions were detected. There were no
    significant differences between groups in respect of weights of liver,
    kidneys, testes and pancreas except that, in females only in the group
    fed at the higher level (0.2%), the weights of the liver, kidneys and
    pancreas were significantly less than those in the control group,
    being about 90% of control; however, the ranges overlapped and the
    means of the test group were well within the range of the control
    group for each organ. Mean values for serum total nitrogen and serum
    urea were presented for females in all three groups and for males in
    the 2 test groups (but not the control group); serum urea was elevated

    to 13% above control in females in the high test group; in all other
    cases the values were below those for the female control group
    (Ferrando, 1971).

    Pig

         European and Landrace pigs of about 50 days of age were divided
    into male control, female control, male test and female test groups,
    each containing 10 pigs; littermates of each sex were divided between
    control and test groups. The feedstuff of test groups contained
    200 000 Anson Units of protease activity per kilogram. The trial
    lasted 119 days. Feed intake, body weight increase and efficacy of
    conversion did not differ significantly between control and test
    groups. The pigs were slaughtered at the end of the trial and
    carcasses, organs and intestine from all the pigs were rated and
    classified as fit for sale by a veterinary surgeon; there were no
    significant differences between control and test groups on the ratings
    of quality (Istituto de Farmacologia, Española, 1978).

    Chicken

         Day-old chicks of Arbon Acres and Vantress types were fed with
    growing mixtures in flour or granule form for 56 days. Test feed-stuff
    contained 800 000 Anson Units of protease activity per kilogram.
    Observations were made on 200 chicks in the laboratory and 13 000
    chicks under farm conditions, each divided into test and control
    groups. At the end of the trial period, mortality was less in the
    treated than in the control farm chicks. No adverse effects were
    observed (Istituto de Farmacologia, Española, 1968).

    Comments

         No adverse effects have been noted in short-term trials with
    S. fradiae protease incorporated in feedstuff at levels (Anson
    Units/kg) of 4 000 000 in rats, 200 000 in pigs, and 800 000 in
    chicks. However, observations were limited to macroscopic post mortem
    examination and serum urea and nitrogen assays in rats and to standard
    meat-inspection procedures in pigs and chicks.

         The level of use of the protease in, for instance, beer appears
    to be about 1000 Anson Units per litre.

         The Committee allocated a temporary ADI "not specified"
    pending the submission of the ongoing feeding studies, including
    histopathological observations, and information about the occurrence
    in nature of the microorganism from which the enzyme is obtained.

    EVALUATION

    Estimate of temporary acceptable daily intake for man

    Not specified.*

    FURTHER WORK OR INFORMATION

    Required by 1984

    (1) Submission of the results of a feeding study, including
    histopathological observations.

    (2) Information about the occurrence of Streptomyces fradiae in
    nature.

              

    *    The statement "ADI not specified" means that, on the basis of the
         available data (toxicological, biochemical, and other), the total
         daily intake of the substance, arising £rom its use or uses at
         the levels necessary to achieve the desired effect and from its
         acceptable background in food, does not, in the opinion of the
         Committee, represent a hazard to health. For this reason, and for
         the reasons stated in individual evaluations, the establishment
         of an acceptable daily intake (ADI) in mg/kg bw is not deemed
         necessary.

    REFERENCES

    Ferrando, R. (1971) Compte rendu des essais de recherche de toxicité
         à moyen terme de la protease "2000" lot or 405 B (2ème essai).
         Appendix D 1 in Appendices to Technical Information on
         Streptomyces fradiae protease manufactured by Compañia Española
         de la Penicilina y Antibioticos, S.A., Madrid, Spain. Technical
         Report No. 2, 10 May 1979. Unpublished report submitted to WHO

    Istituto de Farmacologia, Española (1968) Tests on chicks. Appendix D
         3 in Appendices to Technical Information on Streptomyces
         fradiae protease manufactured by Compañia Española de la
         Penicilina Antibioticos, S.A., Madrid, Spain. Technical Report
         No. 2, 10 May 1979. Unpublished report submitted to WHO

    Istituto de Farmacologia, Española (1978) Estudio de Fradiase en
         crecimiento de ganado porcino en cria industrial. Appendix D 2 in
         Appendices to Technical Information on Streptomyces fradiae
         protease manufactured by Compañia Española de la Penicilina
         Antibioticos, S.A., Madrid, Spain. Technical Report No. 2, 10 May
         1979. Unpublished report submitted to WHO
    


    See Also:
       Toxicological Abbreviations